Department of Biochemistry, Microbiology & Immunology, College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
Department of Biochemistry, Microbiology & Immunology, College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
J Biol Chem. 2022 May;298(5):101951. doi: 10.1016/j.jbc.2022.101951. Epub 2022 Apr 18.
Linear replicons can be found in a minority of prokaryotic organisms, including Borrelia species and Agrobacterium tumefaciens. The problem with replicating the lagging strand end of linear DNAs is circumvented in these organisms by the presence of covalently closed DNA hairpin telomeres at the DNA termini. Telomere resolvases are enzymes responsible for generating these hairpin telomeres from a dimeric replication intermediate through a two-step DNA cleavage and rejoining reaction referred to as telomere resolution. It was previously shown that the agrobacterial telomere resolvase, TelA, possesses ssDNA annealing activity in addition to telomere resolution activity. The annealing activity derives, chiefly, from the N-terminal domain. This domain is dispensable for telomere resolution. In this study, we used activity analyses of an N-terminal domain deletion mutant, domain add back experiments, and protein-protein interaction studies and we report that the N-terminal domain of TelA is involved in inhibitory interactions with the remainder of TelA that are relieved by the binding of divalent metal ions. We also found that the regulation of telomere resolution by the N-terminal domain of TelA extends to suppression of inappropriate enzymatic activity, including hairpin telomere fusion (reaction reversal) and recombination between replicated telomeres to form a Holliday junction.
线性复制子存在于少数原核生物中，包括伯氏疏螺旋体和根癌农杆菌。在这些生物体中，线性 DNA 的滞后链末端的复制问题通过 DNA 末端共价闭合的 DNA 发夹端粒来解决。端粒酶是负责通过两步 DNA 切割和连接反应（称为端粒解决）从二聚体复制中间体产生这些发夹端粒的酶。先前已经表明，农杆菌端粒酶 TelA 除了具有端粒解决活性外，还具有 ssDNA 退火活性。退火活性主要来自 N 端结构域。该结构域对于端粒解决不是必需的。在这项研究中，我们使用了 N 端结构域缺失突变体的活性分析、结构域添加回实验以及蛋白质-蛋白质相互作用研究，并报告称 TelA 的 N 端结构域参与与 TelA 其余部分的抑制相互作用，这些相互作用通过二价金属离子的结合得到缓解。我们还发现，TelA 的 N 端结构域对端粒解决的调节作用扩展到抑制不适当的酶活性，包括发夹端粒融合（反应逆转）和复制端粒之间的重组形成霍利迪连接。
